Field of the Invention
The invention relates to the preparation of thin tissue sections for mass spectrometric imaging with ionization by matrix-assisted laser desorption (MALDI), particularly for high-resolution and high-sensitivity imaging of soluble peptides and proteins.
Description of the Related Art
Imaging mass spectrometry (imaging MS or IMS) acquires images of thin tissue sections which shows for every pixel the mass spectrum of the ionizable components, corresponding to the color spectrum in every pixel of an optical image. The ionization is usually carried out by matrix-assisted laser desorption (MALDI), not least because MALDI primarily produces singly charged molecular ions, which means that the resulting mass spectra are very easy to interpret.
Several methods are known for the preparation of thin tissue sections in imaging mass spectrometry with MALDI, particularly spray methods for matrix solutions. For example, the patent family U.S. Pat. No. 7,667,196 B2, GB 2437623 B and DE 10 2006 019 530 B4 (M. Schürenberg et al.) discloses a method of applying the matrix by depositing isolated droplets of a thin mist of matrix solution, with intermediate drying phases. The individual droplets each bring solvent, which penetrates into the thin tissue section and, via the subsequent vaporization, transports analyte molecules from the thin tissue section to the matrix layer. Unfortunately, the liquid of each droplet spreads radially in all directions, so the analyte substances are also transported laterally within the thin tissue section; this limits the spatial resolving power of the image because the spatial information of the analyte molecules is smeared out.
It is furthermore known that the matrix material can also be applied by resublimation, see for example “Sublimation as a Method of Matrix Application for Mass Spectrometric Imaging”, J. A. Hankin et al., J. Am. Soc. Mass Spectrom. 2007, 18/9, 1646-1652. This method is particularly suitable for phospholipids because their outstandingly good ionizability means that they always display a high sensitivity. The sensitivity for peptides is low, however, because hardly any peptide molecules are embedded into the matrix layer. Only analyte molecules from the topmost molecular layer of the thin tissue section can be captured because the dry application of the matrix layer does not involve a transport of analyte substances from the thin layer to the matrix layer. The consequence of this is that only a small number of analyte molecules are formed during laser bombardment in the course of an ionization with matrix-assisted laser desorption, and the signal-to-noise ratio is unfavorably low at the ion detector.
It is also known that, in a moisture chamber, the swelling of a thin tissue section with a matrix layer applied in the dry state causes small quantities of peptides and proteins to be transferred by diffusion from the volume of the thin tissue section close to the surface into the matrix layer. However, this slow diffusion of the analyte molecules to the matrix layer in the swollen thin tissue section is also associated with an undesirable lateral diffusion of the analyte molecules in the same order of magnitude, which in turn smears out the spatial information of the analyte molecules.
In view of the foregoing, there is a need for a method whereby larger quantities of soluble analyte molecules are transported from the depth of the thin tissue section (i.e. not only from the layers in the immediate vicinity of the surface) into the matrix layer, which may already exist or may be growing at the same time, substantially without lateral diffusion of the analyte molecules.